Compounds having cytokine inhibitory activity

ABSTRACT

There are disclosed compounds of formula (I)and pharmaceutically acceptable salts thereof which exhibit utility for the treatment of cytokine mediated diseases such as arthritis.

This application claims the benefit of Provisional application No.60/142,002, filed Jul. 7, 1999.

BACKGROUND OF THE INVENTION

The present invention relates to substituted imidazole compounds whichhave cytokine inhibitory activity. Cytokine mediated diseases andcytokine inhibition, suppression and antagonism are used in the contextof diseases or conditions in which excessive or unregulated productionor activity of one or more cytokines occurs. Examples of cytokines whichare effected typically include Interleukin-1 (IL-1), Interleukin-6(IL6), Interleukin-8 (IL-8) and Tumor Necrosis Factor (TNF).

Interleukin-1 (IL-1) and Tumor Necrosis Factor (TNF) are produced by avariety of cells which are involved in immunoregulation and otherphysiological conditions.

There are many disease states in which IL-1 is implicated. Examples arerheumatoid arthritis, osteoarthritis, endotoxemia, toxic shock syndrome,acute and chronic inflammatory diseases, such as the inflammatoryreaction induced by endotoxin or inflammatory bowel disease;tuberculosis, atherosclerosis, muscle degeneration, cachexia, psoriaticarthritis, Reiter's syndrome, rheumatoid arthritis, gout, traumaticarthritis, rubella arthritis and acute synovitis. Recent evidence alsolinks IL-1 activity to diabetes.

Interleukin-1 has been demonstrated to mediate a variety of biologicalactivities thought to be important in immunoregulation and otherphysiological conditions. [See, e.g., Dinarello et al., Rev. Infect.Disease, 6, 51 (1984)]. The known biological activities of IL-1 includethe activation of T helper cells, induction of fever, stimulation ofprosta-glandin or collagenase production, neutrophil chemotaxis,induction of acute phase proteins and the suppression of plasma ironlevels.

Excessive or unregulated tumor necrosis factor (TNF) production oractivity has been implicated in mediating or exacerbating rheumatoidarthritis, rheumatoid spondylitis, osteoarthritis, gouty arthritis, andother arthritic conditions, sepsis, septic shock, endotoxic shock, gramnegative sepsis, toxic shock syndrome, adult respiratory distresssyndrome, cerebral malaria, chronic pulmonary inflammatory disease,silicosis, pulmonary sarcosis, bone resorption diseases, reperfusioninjury, graft v. host rejection, allograft rejections, fever and myalgiadue to infection, cachexia secondary to infection or malignancy,cachexia secondary to acquired immune deficiency syndrome (AIDS), AIDSrelated complex (ARC), keloid formation, scar tissue formation, Crohn'sdisease, ulcerative colitis and pyresis.

Monokines, such as TNF, have also been shown to activate HIV replicationin monocytes and/or macrophages [See Poli, et al., Proc. Natl. Acad.Sci., 87:782-784 (1990)], therefore, inhibition of monokine productionor activity aids in limiting HIV progression. TNF has been implicated invarious roles with other viral infections, such as the cytomegalovirus(CMV), influenza virus and the herpes virus.

Interleukin-6 (IL-6) is a cytokine effecting the immune system andhematopoiesis. It is produced by several mammalian cell types inresponse to agents such as IL-1, and is correlated with disease statessuch as angiofollicular lymphoid hyperplasia.

Interleukin-8 (IL-8) is a chemotactic factor first identified andcharacterized in 1987. Many different names have been applied to IL-8,such as neutrophil attractant/activation protein-1 (NAP-1), monocytederived neutrophil chemotactic factor (MDNCF), neutrophil activatingfactor (NAF), and T-cell lymphocyte chemotactic factor. Like IL-1, IL-8is produced by several cell types, including mononuclear cells,fibroblasts, endothelial cells and ketainocytes. Its production isinduced by IL-1, TNF and by lipopolysaccharide (LPS). IL-8 stimulates anumber of cellular functions in vitro. It is a chemoattractant forneutrophils, T-lymphocytes and basophils. It induces histamine releasefrom basophils. It causes lysozomal enzyme release and respiratory burstfrom neutrophils, and it has been shown to increase the surfaceexpression of Mac-1 (CD11b/CD 18) on neutrophils without de novo proteinsynthesis.

There remains a need for compounds which are useful in treating cytokinemediated diseases, and as such, inhibit, suppress or antagonize theproduction or activity of cytokines such as IL-1, IL-6, IL-8 and TNF.

SUMMARY OF THE INVENTION

The present invention relates to compound I of the formula

wherein

X is C₁-C₆ alkyl, said alkyl group optionally substituted by 1-3 groupsselected from halogen, hydroxy, CF₃, NH₂ and NO₂; or a heterocyclicgroup connected to the imidazole ring by a direct bond or by C₁-C₆alkyl;

R₁ and R₂ are independently hydrogen or C₁-C₆ alkyl, said alkyl groupoptionally substituted by 1-3 groups selected from halogen, hydroxy,CF₃, NH₂ and NO₂; or

R₁ and R₂ taken together with the nitrogen atom represent an optionallysubstituted 4 to 10 membered non-aromatic heterocyclic ring containingat least one N atom, and optionally containing 1-2 additional N atomsand 0-1 O or S atom; or

R₂ and X taken together represent an optionally substituted 4 to 10membered non-aromatic heterocyclic ring containing at least one N atom,and optionally containing 1-2 additional N atoms and 0-1 O or S atoms;

Q is CH or N;

R₃ is hydrogen or NH(C₁-C₆ alkyl)aryl;

R₄, R₅ and R₆ independently represent a member selected from the groupconsisting of hydrogen, halo, hydroxy, CF₃, NH₂, NO₂, C₁-C₆ alkyl, saidalkyl group optionally substituted by 1-3 groups selected from halogen,hydroxy, CF₃, NH₂ and NO₂, C₁-C₆ alkoxy, said alkoxy group optionallysubstituted by 1-3 groups selected from halogen, hydroxy, CF₃, NH₂ andNO₂; C₃-C₈ cycloalkyl, said cycloalkyl group optionally substituted by1-3 groups selected from halogen, hydroxy, CF₃, NH₂ and NO₂ or aryl,said aryl group optionally substituted by 1-3 groups selected fromhalogen, hydroxy, CF₃, NH₂ and NO₂;

R₇ is hydrogen or C₁-C₆ alkyl, said alkyl group optionally substitutedby 1-3 groups selected from halogen, hydroxy, CF₃, NH₂ and NO₂;

or a pharmaceutically acceptable addition salt and/or hydrate thereof,or where applicable, a geometric or optical isomer or racemic mixturethereof.

This invention also relates to a pharmaceutical composition which iscomprised of a compound of formula I as defined above in combinationwith a pharmaceutically acceptable carrier.

Also included in the invention is a method of treating a cytokinemediated disease in a mammal, comprising administering to a mammalianpatient in need of such treatment an amount of a compound of formula Iwhich is effective for treating said cytokine mediated disease.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to compound I of the formula

wherein

X is C₁-C₆ alkyl, said alkyl group optionally substituted by 1-3 groupsselected from halogen, hydroxy, CF₃, NH₂ and NO₂; or a heterocyclicgroup connected to the imidazole ring by a direct bond or by C₁-C₆alkyl;

R₁ and R₂ are independently hydrogen or C₁-C₆ alkyl, said alkyl groupoptionally substituted by 1-3 groups selected from halogen, hydroxy,CF₃, NH₂ and NO₂; or

R₁ and R₂ taken together with the nitrogen atom represent an optionallysubstituted 4 to 10 membered non-aromatic heterocyclic ring containingat least one N atom, and optionally containing 1-2 additional N atomsand 0-1 O or S atom; or

R₂ and X taken together represent an optionally substituted 4 to 10membered non-aromatic heterocyclic ring containing at least one N atom,and optionally containing 1-2 additional N atoms and 0-1 O or S atoms;

Q is CH or N;

R₃ is hydrogen or NH(C₁-C₆ alkyl)aryl;

R₄, R₅ and R₆ independently represent a member selected from the groupconsisting of hydrogen, halo, hydroxy, CF₃, NH₂, NO₂, C₁-C₆ alkyl, saidalkyl group optionally substituted by 1-3 groups selected from halogen,hydroxy, CF₃, NH₂ and NO₂, C₁-C₆ alkoxy, said alkoxy group optionallysubstituted by 1-3 groups selected from halogen, hydroxy, CF₃, NH₂ andNO₂; C₃-C₈ cycloalkyl, said cycloalkyl group optionally substituted by1-3 groups selected from halogen, hydroxy, CF₃, NH₂ and NO₂ or aryl,said aryl group optionally substituted by 1-3 groups selected fromhalogen, hydroxy, CF₃, NH₂ and NO₂;

R₇ is hydrogen or C₁-C₆ alkyl, said alkyl group optionally substitutedby 1-3 groups selected from halogen, hydroxy, CF₃, NH₂ and NO₂;

or a pharmaceutically acceptable addition salt and/or hydrate thereof,or where applicable, a geometric or optical isomer or racemic mixturethereof.

This invention also relates to a pharmaceutical composition which iscomprised of a compound of formula I as defined above in combinationwith a pharmaceutically acceptable carrier.

Also included in the invention is a method of treating a cytokinemediated disease in a mammal, comprising administering to a mammalianpatient in need of such treatment an amount of a compound of formula Iwhich is effective for treating said cytokine mediated disease.

In a preferred embodiment, there is disclosed a compound of the formula

or a pharmaceutically acceptable salt thereof, wherein:

Q is CH;

X is C₁-C₆ alkyl; or

R₂ and X taken together represent an azetidine group;

R₄, R₅ and R₆ are independently hydrogen or CF₃;

R₇ is CH₃;

R₁ and R₂ are independently hydrogen or C₁-C₆ alkyl; or

R₁ and R₂ taken together with the nitrogen atom form a morpholine group.

Representative species falling within the present invention include thefollowing:

Unless otherwise stated or indicated, the following definitions shallapply throughout the specification and claims.

The term “alkyl” refers to a monovalent alkane (hydrocarbon) derivedradical containing from 1 to 15 carbon atoms unless otherwise defined.It may be straight or branched, and when of sufficient size, e.g., C3-15may be cyclic. Preferred straight or branched alkyl groups includemethyl, ethyl, propyl, isopropyl, butyl and t-butyl. Preferredcycloalkyl groups include cyclopropyl, cyclopentyl and cyclohexyl.

Alkyl also includes an alkyl group substituted with a cycloalkyl group,such as cyclopropylmethyl. Alkyl also includes a straight or branchedalkyl group

The alkylene and monovalent alkyl portion(s) of the alkyl group can beattached at any available point of attachment to the cycloalkyleneportion.

When substituted alkyl is present, this refers to a straight, branchedor cyclic alkyl group as defined above, substituted with 1-3 groups asdefined with respect to each variable.

The term “aryl” refers to aromatic rings e.g., phenyl, substitutedphenyl and like groups as well as rings which are fused, e.g., naphthyland the like. Aryl thus contains at least one ring having at least 6atoms, with up to two such rings being present, containing up to 10atoms therein, with alternating (resonating) double bonds betweenadjacent carbon atoms. The preferred aryl groups are phenyl andnaphthyl. Aryl groups may likewise be substituted as defined below.Preferred substituted aryls include phenyl or naphthyl substituted withone or two groups.

The terms “heterocycloalkyl” and “heterocyclyl” refer to a cycloalkylgroup (nonaromatic) in which one of the carbon atoms in the ring isreplaced by a heteroatom selected from O, S(O)y or N, and in which up tothree additional carbon atoms may be replaced by said heteroatoms. Whenthree heteroatoms are present in the heterocycle, they are not alllinked together.

Examples of heterocyclyls are piperidinyl, morpholinyl, azetidinyl,pyrrolidinyl, tetrahydrofuranyl, imidazolinyl, piperazinyl,pyrolidin-2-one, piperidin-2-one and the like.

The term “halogen” or “halo” is intended to include fluorine, chlorine,bromine and iodine.

As used herein, the term “composition” is intended to encompass aproduct comprising the specified ingredients in the specified amounts,as well as any product which results, directly or indirectly, fromcombination of the specified ingredients in the specified amounts.

In addition, it is well known to those skilled in the art that many ofthe foregoing heterocyclic groups can exist in more than one tautomericform. It is intended that all such tautomers be included within theambit of this invention.

The optical isomeric forms, that is mixtures of enantiomers, e.g.,racemates, or diastereomers as well as individual enantiomers ordiastereomers of the instant compound are included. These individualenantiomers are commonly designated according to the optical rotationthey effect by the symbols (+) and (−), (L) and (D), (1) and (d) orcombinations thereof. These isomers may also be designated according totheir absolute spatial configuration by (S) and (R), which stands forsinister and rectus, respectively.

The individual optical isomers may be prepared using conventionalresolution procedures, e.g., treatment with an appropriate opticallyactive acid, separating the diastereomers and then recovering thedesired isomer. In addition, the individual optical isomers may beprepared by asymmetric synthesis.

Additionally, a given chemical formula or name shall encompasspharmaceutically acceptable addition salts thereof and solvates thereof,such as hydrates.

The compounds of the present invention, while effective themselves, maybe formulated and administered in the form of their pharmaceuticallyacceptable addition salts for purposes of stability, convenience ofcrystallization, increased solubility and other desirable properties.

The compounds of the present invention may be administered in the formof pharmaceutically acceptable salts. The term “pharmaceuticallyacceptable salt” is intended to include all acceptable salts. Examplesof acid salts are hydrochloric, nitric, sulfuric, phosphoric, formic,acetic, trifluoroacetic, propionic, maleic, succinic, malonic, methanesulfonic and the like which can be used as a dosage form for modifyingthe solubility or hydrolysis characteristics or can be used in sustainedrelease or prodrug formulations. Depending on the particularfunctionality of the compound of the present invention, pharmaceuticallyacceptable salts of the compounds of this invention include those formedfrom cations such as sodium, potassium, aluminum, calcium, lithium,magnesium, zinc, and from bases such as ammonia, ethylenediamine,N-methyl-glutamine, lysine, arginine, ornithine, choline,N,N′-dibenzylethylenediamine, chloroprocaine, diethanolamine, procaine,N-benzylphenethylamine, diethylamine, piperazine,tris(hydroxymethyl)aminomethane, and tetramethyl-ammonium hydroxide.These salts may be prepared by standard procedures, e.g. by reacting afree acid with a suitable organic or inorganic base, or alternatively byreacting a free base with a suitable organic or inorganic acid.

Also, in the case of an acid (—COOH) or alcohol group being present,pharmaceutically acceptable esters can be employed, e.g. methyl, ethyl,butyl, acetate, maleate, pivaloyloxymethyl, and the like, and thoseesters known in the art for modifying solubility or hydrolysischaracteristics for use as sustained release or prodrug formulations.

The compounds of the present invention may have chiral centers otherthan those centers whose stereochemistry is depicted in formula I, andtherefore may occur as racemates, racemic mixtures and as individualenantiomers or diastereomers, with all such isomeric forms beingincluded in the present invention as well as mixtures thereof.Furthermore, some of the crystalline forms for compounds of the presentinvention may exist as polymorphs and as such are intended to beincluded in the present invention. In addition, some of the compounds ofthe instant invention may form solvates with water or common organicsolvents. Such solvates are encompassed within the scope of thisinvention.

The term “TNF mediated disease or disease state” refers to diseasestates in which TNF plays a role, either by production or increasedactivity levels of TNF itself, or by causing another cytokine to bereleased, such as but not limited to IL-1 or IL-6. A disease state inwhich IL-1, for instance is a major component, and whose production oraction, is exacerbated or secreted in response to TNF, would thereforebe considered a disease state mediated by TNF.

The term “cytokine” as used herein means any secreted polypeptide thataffects the functions of cells and is a molecule which modulatesinteractions between cells in the immune, inflammatory or hematopoieticresponse. A cytokine includes, but is not limited to, monokines andlymphokines regardless of which cells produce them. Examples ofcytokines include, but are not limited to, Interleukin-1 (IL-1),Interleukin-6 (IL-6), Interleukin-8 (IL-8), Tumor Necrosis Factor-alpha(TNF-α) and Tumor Necrosis Factor-beta (TNF-β).

By the term “cytokine interfering or cytokine suppresive amount” ismeant an effective amount of a compound of formula I which will cause adecrease in the in vivo activity or level of the cytokine to normal orsub-normal levels, when given to the patient for the prophylaxis ortherapeutic treatment of a disease state which is exacerbated by, orcaused by, excessive or unregulated cytokine production or activity.

The compounds of the invention are prepared by the following generalreaction scheme. All substituents are as defined above unless indicatedotherwise. The preparation of specific compounds is illustrated in theexamples section.

The compounds of formula 1 can be used in the prophylactic ortherapeutic treatment of disease states in mammals which are exacerbatedor caused by excessive or unregulated cytokines, e.g., IL-1, IL-6, IL-8or TNF.

Because the compounds of formula I inhibit cytokines, the compounds areuseful for treating diseases in which cytokine presence or activity isimplicated, such as rheumatoid arthritis, rheumatoid spondylitis,osteoarthritis, gouty arthritis and other arthritic conditions.

The compounds of formula I are useful to treat disease states mediatedby excessive or unregulated TNF production or activity. Such diseasesinclude, but are not limited to sepsis, septic shock, endotoxic shock,gram negative sepsis, toxic shock syndrome, adult respiratory distresssyndrome, cerebral malaria, chronic pulmonary inflammatory disease,silicosis, pulmonary sarcoidosis, bone resorption diseases, such asosteoporosis, reperfusion injury, graft v. host rejection, allograftrejection, fever, myalgia due to infection, cachexia secondary toinfection or malignancy, cachexia secondary to acquired immunedeficiency syndrome (AIDS), AIDS, ARC (AIDS related complex), keloidformation, scar tissue formation, Crohn's disease, ulcerative colitis,pyresis, AIDS and other viral infections, such as cytomegalovirus (CMV),influenza virus, and the herpes family of viruses such as Herpes Zosteror Simplex I and II.

The compounds of formula I are also useful topically in the treatment ofinflammation such as in the treatment of rheumatoid arthritis,rheumatoid spondylitis, osteoarthritis, gouty arthritis and otherarthritic conditions; inflamed joints, eczema, psoriasis or otherinflammatory skin conditions such as sunburn; inflammatory eyeconditions including conjunctivitis; pyresis, pain and other conditionsassociated with inflammation.

The compounds of formula I are also useful in treating diseasescharacterized by excessive IL-8 activity. These disease states includepsoriasis, inflammatory bowel disease, asthma, cardiac and renalreperfusion injury, adult respiratory distress syndrome, thrombosis andglomerulonephritis.

The invention thus includes a method of treating psoriasis, inflammatorybowel disease, asthma, cardiac and renal reperfusion injury, adultrespiratory distress syndrome, thrombosis and glomerulonephritis, in amammal in need of such treatment, which comprises administering to saidmammal a compound of formula I in an amount which is effective fortreating said disease or condition.

When administered to a patient for the treatment of a disease in which acytokine or cytokines are implicated, the dosage used can be variedwithin wide limits, depending upon the type of disease, the age andgeneral condition of the patient, the particular compound administered,the presence or level of toxicity or adverse effects experienced withthe drug and other factors. A representative example of a suitabledosage range is from as low as about 0.01 mg/kg to as high as about 100mg/kg. However, the dosage administered is generally left to thediscretion of the physician.

The methods of treatment can be carried out by delivering the compoundof formula I parenterally. The term ‘parenteral’ as used herein includesintravenous, intramuscular, or intraperitoneal administration. Thesubcutaneous and intramuscular forms of parenteral administration aregenerally preferred. The instant invention can also be carried out bydelivering the compound of formula I subcutaneously, intranasally,intrarectally, transdermally or intravaginally.

The compounds of formula I may also be administered by inhalation. By‘inhalation’ is meant intranasal and oral inhalation administration.Appropriate dosage forms for such administration, such as an aerosolformulation or a metered dose inhaler, may be prepared by conventiontechniques.

The invention also relates to a pharmaceutical composition comprising acompound of formula I and a pharmaceutically acceptable carrier. Thecompounds of formula I may also be included in pharmaceuticalcompositions in combination with a second therapeutically activecompound.

The pharmaceutical carrier employed may be, for example, either a solid,liquid or gas. Exemples of solid carriers include lactose, terra alba,sucrose, talc, gelatin, agar, pectin, acacia, magnesium stearate,stearic acid and the like. Exemples of liquid carriers are syrup, peanutoil, olive oil, water and the like. Examples of gaseous carriers includecarbon dioxide and nitrogen.

Similarly, the carrier or diluent may include time delay material wellknown in the art, such as glyceryl monostearate or glyceryl distearate,alone or with a wax.

A wide variety of pharmaceutical dosage forms can be employed. If asolid dosage is used for oral administration, the preparation can be inthe form of a tablet, hard gelatin capsule, troche or lozenge. Theamount of solid carrier will vary widely, but generally will be fromabout 0.025 mg to about 1 g. When a liquid dosage form is desired fororal administration, the preparation is typically in the form of asyrup, emulsion, soft gelatin capsule, suspension or solution. When aparenteral dosage form is to be employed, the drug may be in solid orliquid form, and may be formulated for administration directly or may besuitable for reconstitution.

Topical dosage forms are also included. Examples of topical dosage formsare solids, liquids and semi-solids. Solids would include dustingpowders, poultices and the like. Liquids include solutions, suspensionsand emulsions. Semi-solids include creams, ointments, gels and the like.

The amount of a compound of formula I used topically will, of course,vary with the compound chosen, the nature and severity of the condition,and can be varied in accordance with the discretion of the physician. Arepresentative, topical, dose of a compound of formula I is from as lowas about 0.01 mg to as high as about 2.0 g, administered one to four,preferably one to two times daily.

The active ingredient may comprise, for topical administration, fromabout 0.001% to about 10% w/w.

Drops according to the present invention may comprise sterile ornon-sterile aqueous or oil solutions or suspensions, and may be preparedby dissolving the active ingredient in a suitable aqueous solution,optionally including a bactericidal and/or fungicidal agent and/or anyother suitable preservative, and optionally including a surface activeagent. The resulting solution may then be clarified by filtration,transferred to a suitable container which is then sealed and sterilizedby autoclaving or maintaining at 98-100° C. for half an hour.Alternatively, the solution may be sterilized by filtration andtransferred to the container aseptically. Examples of bactericidal andfungicidal agents suitable for inclusion in the drops are phenylmercuricnitrate or acetate (0.002%), benzalkonium chloride (0.01%) andchlorhexidine acetate (0.01%). Suitable solvents for the preparation ofan oily solution include glycerol, diluted alcohol and propylene glycol.

Lotions according to the present invention include those suitable forapplication to the skin or eye. An eye lotion may comprise a sterileaqueous solution optionally containing a bactericide and may be preparedby methods similar to those for the preparation of drops. Lotions orliniments for application to the skin may also include an agent tohasten drying and to cool the skin, such as an alcohol or acetone,and/or a moisturizer such as glycerol or an oil such as castor oil orarachis oil.

Creams, ointments or pastes according to the present invention aresemi-solid formulations of the active ingredient for externalapplication. They may be made by mixing the active ingredient infinely-divided or powdered form, alone or in solution or suspension inan aqueous or non-aqueous liquid, with a greasy or non-greasy base. Thebase may comprise hydrocarbons such as hard, soft or liquid paraffin,glycerol, beeswax, a metallic soap; a mucilage; an oil of natural originsuch as almond, corn, arachis, castor or olive oil; wool fat or itsderivatives, or a fatty acid such as stearic or oleic acid together withan alcohol such as propylene glycol or macrogels. The formulation mayincorporate any suitable surface active agent such as an anionic,cationic or non-ionic surfactant such as sorbitan esters orpolyoxyethylene derivatives thereof. Suspending agents such as naturalgums, cellulose derivatives or inorganic materials such as silicas, andother ingredients such as lanolin may also be included.

The following examples illustrate the preparation of some of thecompounds of the invention and are not to be construed as limiting theinvention disclosed herein.

EXAMPLE 1

(S)-1-phenyl-N-{4-[2-(2-amino-ethyl)-3-methyl-5-(3-trifluoromethyl-phenyl)-3H-imidazol-4-yl]-pyridin-2-y}-ethylamine

Step 1A

2-(2-Fluoropyridin-4-yl)-1-(3-trifluoromethylphenyl)-ethanone

To a suspension of N,O-dimethylhydroxylamine hydrochloride (58.2 g, 0.60mol) in dichloromethane (1 L) at 0° C., under argon, was added3-trifluoromethylbenzoyl chloride (104.0 g, 0.50 mol), followed by aslow addition of triethylamine (152.3 mL, 1.09 mol). The reaction wasaged for 30 min. at 5° C. and then allowed to warm to room temperature.TLC (1:1, ethyl acetate/hexane) showed the reaction to be complete. Thereaction was washed with 5% aqueous citric acid (500 mL) and 5% aqueoussodium bicarbonate. The aqueous extracts were back extracted withmethylene chloride (100 mL) and the combined methylene chloride extractswere dried over sodium sulfate, filtered and concentrated to an oil. Theoil was redissolved in toluene (2×100 mL) and evaporated in vacuo toafford the Weinreb amide (114.7 g).

¹H NMR (300 MHz, CDCl3) d 7.98 (s, 1H, Ar), 7.89 (d, J=7.8 Hz, 1H, Ar),7.72 (d, J=7.8 Hz, 1H, Ar), 7.55 (t, J=7.8 Hz, 1H, Ar), 3.55 (s, 3H,CH₃O), 3.39 (s, 3H, CH₃N).

To a stirring solution of diisopropylamine (17.69 mL, 0.135 mol) inanhydrous THF (200 mL) at −78° C., under argon, was added n-butyllithium(54.0 mL, 2.5M in hexane, 0.135 mol), followed after 5 min. by asolution of 2-fluoro-4-methylpyridine (10 g, 0.090 mol) in anhydrous THF(20 mL). After stirring for 15 min. at −78° C., a solution ofN-methoxy-N-methyl-3-trifluoromethylbenzamide (23.08 g, 0.099 mol) inanhydrous THF (10 mL) was added to the reaction mixture which was thenstirred for 5 min., and allowed to warm to 0° C. The reaction wasquenched with water (400 mL), and extracted with ethyl acetate (3×200mL). The ethyl acetate extracts were combined, dried over anhydroussodium sulfate, filtered, and concentrated to an oil which waschromatographed on silica gel (1 kg), eluting with 20% ethyl acetate inhexane to give 21.6 g of the title compound.

¹H NMR (300 MHz, CDCl₃) d 8.25 (s, 1H, Pyr), 8.20 (d, J=5.1 Hz, 1H,Pyr), 8.18 (d, J=9.3 Hz, 1H, Pyr), 7.88 (d, J=7.8 Hz, Ar), 7.67 (t,J=7.8 Hz, 1H, Ar), 7.09 (d, J=5.1 Hz, 1H, Ar), 6.86 (s, 1H, Ar), 4.37(s, 2H, PyrCH₂C).

Step 1B

1-(2-Fluoropyridin-4-yl)-2-(3-trifluoromethylphenyl)-ethane-1,2-dione1-oxime

To a mixture of2-(2-fluoropyridin-4-yl)-1-(3-trifluoromethylphenyl)ethanone (10.80 g,0.038 mol) in ethanol (200 mL), at −10° C., under argon, was addedtert-butyl nitrite (5.0 mL, 0.042 mol) and hydrogen chloride (12.2 mL,2.5M in ethanol, 0.031 mol) dropwise while maintaining the temperaturebelow −5° C. Upon completion of addition, the reaction was allowed towarm to RT for 2 hrs. The reaction mixture was then concentrated invacuo, diluted with water (100 mL), basified with saturated sodiumbicarbonate (200 ml), and extracted with ethyl acetate (3×400 mL). Thecombined organic layers were washed with water (300 mL), dried withbrine (300 mL) and anhydrous sodium sulfate, filtered and concentratedto yield 11.4 g of the title compound, as an oil.

¹H NMR (300 MHz, CDCl₃)d 8.31 (s, 1H, Ar), 8.29 (d, J=5.3 Hz, 1H, Pyr),8.24 (d, J=7.8 Hz, 1H, Ar), 7.92 (d, J=8.1 Hz, 1H, Ar), 7.71 (t, J=7.8Hz, 1H, Ar), 7.40 (d, J=5.1 Hz, 1H, Pyr), 7.23 (s, 1H, Pyr).

Step 1C

2-amino-2-(2-fluoropyridine-4-yl)-1-(3-trifluoromethylphenyl)ethanol

10% Palladium on carbon (3.0 g) was added to a solution of1-(2-fluoropyridin4yl)-2-(3-trifluoromethylphenyl)-ethane-1,2-dione1-oxime (8.0 g, 27 mmol) in ethanol (400 mL) at ambient temperature. Thereaction vessel was vacuum purged, refilled with hydrogen and vigorouslystirred for 10 hrs. After the reaction was complete, the solution wasfiltered through a pad of celite, and concentrated to give a solid. Theresidue was purified by recrystallization from methylene chloride andhexane to give 7.4 g of the title compound. m.p. 128-129° C.

¹H NMR (300 MHz, CD₃OD) d 8.01 (d, J=5.0 Hz, 1H, Ar), 7.53 (m, 1H, Ar),7.49 (m, 2H, Ar), 7.43 (s, 1H, Ar), 7.06 (d, J=5.0 Hz, 1H, Ar), 6.86 (s,1H, Ar), 4.96 (d, J=5.0 Hz, 1H, CH), 4.12 (d, J=5.0 Hz, 1H, CH).

Step 1D

2-(2-fluoropyridin-4-yl)-2-methylamino-1-(3-trifluoromethyl-phenyl)-ethanol

2-Amino-2-(2-fluoropyridin-4-yl)-1-(3-trifluoromethyl-phenyl)-ethanol (6g, 20 mmol) was dissolved in ethyl formate (80 mL) and heated to refluxfor 10 hrs under an argon atmosphere. The reaction mixture was cooled toambient temperature and concentrated to yield the formamide as an oil.IH NMR (300 MH, CD₃0D) d 8.06 (m, 2 H, Ar), 7.55 (m, 3 H, Ar), 7.05 (m,1 H,

¹H NMR (300 MHz, CD₃OD) d 8.06 (m, 2H, Ar), 7.55 (m, 3H, Ar), 7.05 (m,1H, Ar), 5.22 (d, J=5.7 Hz, 1H, CH), 5.10 (d, J=5.7 Hz, 1H, CH).

The formamide was azeotroped with toluene (2×100 mL) to remove traceamounts of ethyl formate and water. Reduction to the desired compoundwas carried out by drop-wise addition of borane-tetrahydrofuran complex(60 mL of a 1.0 M solution in THF, 60 mmol) to a solution the formamideproduct (20 mmol) in THF (60 mL) at ambient temperature. After 1 hr, thereaction was quenched by slow addition of the organic mixture to avigorously stirring solution of aqueous hydrochloric acid (2.0 M, 500mL). Stirring was continued for 3 h to assure complete dissociation ofboron complexes. A solution of aqueous sodium hydroxide (10 M) was thenadded until a pH of 11 was achieved. The resulting mixture was extractedwith ethyl acetate (2×300 mL), dried (sodium sulfate), and concentrated.The resulting residue was purified by filtration through a pad of silicagel (using 5% methanol and 0.5% ammonium hydroxide mixture in methylenechloride) to yield 5.3 g of a solid: mp 99-102° C.

¹H NMR (300 MHz, CD₃OD) d 7.99 (d, J=5.1 Hz, 1H, Ar), 7.50 (m, 1H, Ar),7.45 (m, 2H, Ar), 7.38 (s, 1H, Ar), 7.02 (m, 1H, Ar), 6.83 (s, 1H, Ar),5.09 (d, J=4.8 Hz, 1H, CH), 3.85 (d, J=4.8 Hz, 1H, CH), 2.22 (s, 3H,CH₃).

Step 1E

3-(Benzyloxycarbonyl-amino)-N-[1-(2-fluoro-pyridin-4-yl)-2-hydroxy-2-(3-trifluoromethyl-phenyl)-ethyl]-N-methyl-propionamide

1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (0.61 g, 3.2mmol) was added to a solution of2-(2-fluoro-pyridin-4-yl)-2-methylamino-1-(3-trifluoromethyl-phenyl)-ethanol(0.50 g, 1.6 mmol), 3-(benzyloxycarbonnyl-amino)-propanoic acid (0.53 g,2.4 mmol), triethylamine (0.89 mL, 6.4 mmol), and1-hydroxy-7-azabenzotriazole (0.43 g, 3.9 mmol) in dimethylformamide (10mL) at ambient temperature. After 2 hrs stirring at ambient temperature,ethyl acetate was added (200 mL) followed by aqueous citric acid (50 mLof a 10% solution). The organic layer was then washed with aqueoussodium bicarbonate (50 mL of a sat. solution) and water (3×30 mL), dried(sodium sulfate), filtered, and concentrated. The crude residue waspassed through a pad of silica gel (50% ethyl acetate in hexane) toremove minor impurities and yield 1.4 g of an oil.

¹H MMR (300 MHz, CD₃OD) d 8.17 (d, J=6.1 Hz, 1H, Pyr), 8.01 (s, 1H, Ar),7.67 (d, J =7.8 Hz, 1H, Ar), 7.61 (d, J=7.8 Hz, 1H, Ar), 7.48 (t, J=7.8Hz, 1H, Ar), 7.42-7.32 (m, 5H, Ar), 7.19 (d, J=7.1 Hz, 1H, Pyr), 7.05(s, 1H, Pyr), 5.61 (d, J=8.8 Hz, 1H, PhCHOH), 5.40 (d, J=8.8 Hz, 1H,PhCHN), 5.03 (s, 2H, PhCH₂O), 3.42-3.25 (m, 4H, HNCH₂CH₂), 2.36 (s, 3H,CH₃N).

Step 1F

N-benzyloxycarbonyl-2-[5-(2-fluoro-pyridin-4-yl)-1-methyl-4-(3-trifluoromethyl-phenyl)-1H-imidazol-2-yl]-ethylamine

Oxalyl chloride (0.34 mL, 3.9 mmol) was added to a solution of dimethylsulfoxide (0.41 mL, 5.8 mmol) in methylene chloride (10 mL) at −78° C.After 20 min. at −78° C.,3-(benzyloxycarbonyl-amino)-N-[1-(2-fluoro-pyridin4-yl)-2-hydroxy-2-(3-trifluoromethyl-phenyl)ethyl]-N-methyl-propionamide(1.0 g, 1.9 mmol) in methylene chloride (10 mL) was added and thereaction solution was stirred at −78° C. for 2 hrs. Triethylamine (1.3mL, 9.6 mmol) was added and the cooling bath was removed. The solutionwas diluted with ethyl acetate (150 mL), washed with aqueous ammoniumchloride (75 mL) and brine (75 mL), dried (sodium sulfate), andconcentrated. Some non-polar impurities were removed by passing theresidue through silica gel, eluting with 50% ethyl acetate in hexane.The ketone was then transferred to a flask containing anhydrous ammoniumtrifluoroacetate (4 g) using a minimum amount of ether-methylenechloride. The mixture was placed under vacuum to remove the solvent, andthen placed in a preheated oil bath (150° C.). Once all solid had meltedand efficient stirring was achieved (approx. 10 min.) the formation ofthe imidazole was found to be complete. The heating bath was removed andthe solids were partitioned between ethyl acetate and water (150/50 mL).The organic layer was dried (sodium sulfate), filtered, andconcentrated. The product was purified by filtration through silica gel(imidazole is the least polar component) using ethyl acetate to yield735 mg of a foam.

¹H NMR (300 MHz, CD₃OD) d 8.29 (d, J=5.4 Hz, 1H, Pyr), 7.69 (s, 1H, Ar),7.53 (d, J=7.8 Hz, 1H, Ar), 7.52 (d, J=7.8 Hz, 1H, Ar), 7.45 (t, J=7.8Hz, 1H, Ar), 7.35-7.22 (m, 6H, Ar), 7.05 (s, 1H, Pyr), 5.03 (s, 2H,PhCH₂O), 3.55 (t, J=6.8 Hz, 2H, HNCH₂CH), 3.51 (s, 3H, CH₃N), 3.03 (t,J=6.7 Hz, 2H, CCH₂CH₂).

Step 1G

(S)-1-phenyl-N-{4-[2-(2-(benzyloxycarbonyl-amino)ethyl)-3-methyl-5-(3-trifluoromethyl-phenyl)-3H-imidazol-4-yl]-pyridin-2-yl}-ethylamine

N-Benzyloxycarbonyl-2-[5-(2-fluoro-pyridin-4-yl)-1-methyl-4-(3-trifluoromethyl-phenyl)-1H-imidazol-2-yl]-ethylamine(0.35 g, 0.70 mmol) was dissolved in S-(−)-a-methylbenzylamine (0.85 g,7.0 mmol) and the reaction mixture was heated to 150° C. for 15 hrs. Theheating bath was removed and the contents of the flask were partitionedbetween ethyl acetate (100 mL) and pH 4.5 buffer (50 mL composed of 10%citric acid that was treated with 10 N sodium hydroxide to achieve a pHof 4.5). The organic layer was dried (sodium sulfate), filtered, andconcentrated. The residue was purified by silica gel chromatography toyield 315 mg of the desired product as an oil.

¹H NMR (300 MHz, CD₃OD) d 7.99 (d, J=5.4 Hz, 1H, Pyr), 7.69 (s, 1H, Ar),7.49 (d, J=8.8 Hz, 1H, Ar), 7.47 (d, J=9.3 Hz, 1H, Ar), 7.36 (t, J=7.7Hz, 1H, Ar), 7.33-7.13 (m, 10H, Ar), 6.43 (d, J=5.4 Hz, 1H, Pyr), 6.33(s, 1H, Pyr), 5.03 (s, 2H, PhCH₂O), 4.81 (q, J=6.8 Hz, 1H, PhCHCH₃),3.50 (t, J=6.8 Hz, 2H, HNCH₂CH₂), 3.32 (s, 3H, CH₃N), 2.98 (t, J=6.7 Hz,2H, CCH₂CH₂), 1.48 (d, J=6.8 Hz, 3H, PhCHCH₃).

Step 1H

(S)-1-phenyl-N-{4-[2-(2-amino-ethyl)-3-methyl-5-(3-trifluoromethyl-phenyl)-3H-imidazol-4-yl]-pyridin-2-yl}-ethylamine

A solution of(S)-1-phenyl-N-{4-[2-(2-(benzyloxycarbonyl-amino)-ethyl)-3-methyl-5-(3-trifluoromethyl-phenyl)-3H-imidazol-4-yl]-pyridin-2-yl}-ethylamine(0.20 g, 0.33 mmol) in absolute ethanol (10 mL) was flushed with argonbefore 10% palladium on activated carbon (0.07 g) was added. Thisreaction mixture, flushed with hydrogen gas, was vigorously stirredunder one atmosphere of hydrogen for 5 hrs, then filtered andconcentrated. The residue was purified by silica gel chromatography toyield 146 mg of the desired product as an oil.

¹NMR (300 MHz, CD₃OD) d 7.99 (d, J=5.4 Hz, 1H, Pyr), 7.70 (s, 1H, Ar),7.51 (d, J=7.6 Hz, 1H, Ar), 7.46 (d, J=7.8 Hz, 1H, Ar), 7.37 (t, J=7.7Hz, 1H, Ar), 7.32-7.14 (m, 5H, Ar), 6.47 (d, J=5.4 Hz, 1H, Pyr), 6.35(s, 1H, Pyr), 4.81 (q, J=7.1 Hz, 1H, PhCHCH₃), 3.35 (s, 3H, CH₃N), 3.06(t, J=6.7 Hz, 2H, CH₂CH₂NH₂), 2.93 (t, J=7.0 Hz, 2H, CCH₂CH₂), 1.48 (d,J=6.8 Hz, 3H, PhCHCH₃).

EXAMPLE 2

(S)-1-Phenyl-N-{4-[2-((R)-1-amino-ethyl)-3-methyl-5-(3-trifluoromethyl-phenyl)-3H-imidazol-4-yl]-pyridin-2-yl}-ethylamine

The compound was prepared as described in example 1, replacing3-(benzyloxycarbonnyl-amino)-propanoic acid with N-Cbz-D-Alanine in step1E.

¹H NMR (300 MHz, CD₃OD) d 8.00 (d, J=5.4 Hz, 1H, Pyr), 7.72 (s, 1H, Ar),7.51 (d, J=9.1 Hz, 1H, Ar), 7.48 (d, J=9.2 Hz, 1H, Ar), 7.37 (t, J=7.7Hz, 1H, Ar), 7.31-7.22 (m, 4H, Ar), 7.16 (t, J=6.8 Hz, 1H, Ar), 6.46 (m,1H, Pyr), 6.33 (s, 1H, Pyr), 4.80 (q, J=6.3 Hz, 1H, PhCHCH₃), 4.26 (q,J=6.6 Hz, 1H, NH₂CHCH₃), 3.41 (s, 3H, NCH₃), 1.53 (d, J=6.8 Hz, 3H,PhCHCH₃), 1.48 (d, J=6.8 Hz, 3H, NH₂CHCH₃); MS (FAB) Calcd forC₂₆H₂₆F₃N₅ (M+H⁺) 466.2, found 466.3

EXAMPLE 3

(S)-1-Phenyl-N-{4-[2-(azetidin-3-yl-methyl)-3-methyl-5-(3-trifluoromethyl-phenyl)-3H-imidazol-4-yl]-pyridin-2-yl}-ethylamine

Compound was prepared as described in example 1, replacing3-(benzyloxycarbonnyl-amino)-propanoic acid with3-carboxymethyl-azetidine-1-carboxylic acid benzyl ester (synthesisdescribed below) in step E.

¹H NMR (300 MHz, CD₃OD) d 7.99 (d, J=5.1 Hz, 1H, Pyr), 7.67 (s, 1H, Ar),7.49 (d, J=8.3 Hz, 1H, Ar), 7.46 (d, J=9.5 Hz, 1H, Ar), 7.36 (t, J=7.7Hz, 1H, Ar), 7.32−7.13 (m, 5H, Ar), 6.45 (d, J=5.1 Hz, 1H, Pyr), 6.33(s, 1H, Pyr), 4.80 (q, J=6.8 Hz, 1H, PhCHCH₃), 3.94 (m, 2H, CHCH₂NH),3.69 (m, 2H, CHCH₂NH), 3.34 (s, 3H, NCH₃), 3.10 (d, J=7.6 Hz, 2H,CCH₂CH), 3.04 (m, 1H, CH₂CHCH₂), 1.48 (d, J=6.8 Hz, 3H, PhCHCH₃); MS(FAB) Calcd for C₂₈H₂₈F₃N₅ (M+H⁺) 492.2, found 492.3

Step 3A

1-Benzbydryl-azetidin-3-ol Hydrochloride

A mixture of benzhydrylamine (56 g, 306 mmol), and epichlorohydrin (24mL, 306 mmol) in methanol (150 mL) was stirred at 24° C. under argon for3 days, then refluxed for 3 days. This reaction mixture was concentratedin vacuo, the residue stirred in acetone (500 mL) and filtered to givethe crystaline salt as the title compound (78 g).

Step 3B

(1-Benzhydryl-azetidin-3-ylidene)-acetic Acid Ethyl Ester

To a stirring solution of DMSO (5.3 mL, 75 mmol) in methylene chloride(100 mL) at −78° C. under argon was added oxalyl chloride (4.6 mL, 53mmol) dropwise. After stirring for 30 min., a solution of1-benzhydryl-azetidin-3-ol hydrochloride (12) (10 g, 36 mmol) inmethylene chloride (10 mL) was added dropwise. The reaction mixture wasstirred for 1 hr at −78° C. before triethylamine (18 mL, 128 mmol) wasadded. The ice bath was removed and the reaction quenched with saturatedammonium chloride (75 mL) at 0° C., then extracted with ethyl acetate(3×150 mL). The combined ethyl acetate layer was washed with water (100mL), brine (100 mL), dried over anhydrous sodium sulfate, filtered, andconcentrated. The residue was stirred in ethyl acetate and hexane,filtered to give 1-benzhydrylazetidin-3-one (9 g, 33 mmol, 92%). To astirring solution of the resulting 1-benzhydryl-azetidin-3-one (8.3 g,30 mmol) in methylene chloride (120 mL) was added triethylamine (6.4 mL,45 mmol), and carbethoxymethylene triphenylphosphorane (15.9 g, 45mmol). This reaction mixture was stirred under argon at room temperaturefor 2 hrs., then quenched with acetone (1 mL), and concentrated invacuo. The residue was stirred in 20% ethyl acetate in hexane (20 mL) astriphenyl phosphine oxide precipitated. The mixture was then filteredand concentrated. The crude product was chromatographed on silica gel,eluting with 50:40:10 methylene chloride:hexane:ether to give 9.3 g ofthe title compound.

Step 3C

Azetidin-3-yl-acetic Acid Ethyl Ester

To a solution of (1-benzhydrylazetidin-3-ylidene)-acetic acid ethylester (5.0 g, 16 mmol) in ethanol (200 mL) was added palladium hydroxideon carbon (1.5 g). This mixture was vigorously shaken under a 50 psihydrogen atmosphere for 24 hrs, then filtered and concentrated to givean oil, used without additional purification.

Step 3D

3-Ethoxycarbonylmethyl-azetidine-1-carboxylic Acid Benzyl Ester

To a stirring mixture of azetidin-3-yl-acetic acid ethyl ester (1.0 g,6.98 mmol), in saturated aqueous sodium bicarbonate (5 mL) and THF (5mL) at 0° C. was added carbobenzyloxy chloride (1.1 mL, 7.68 mmol)dropwise. The ice bath was removed and the reaction mixture was stirredat room temperature for 2 hrs, then extracted with ethyl acetate acetate(3×20 mL). The combined ethyl acetate layer was washed with water (10mL), brine (10 mL), dried over anhydrous sodium sulfate, filtered, andconcentrated. The residue was chromatographed on silica gel, elutingwith 20% ethyl acetate in hexane to give 0.30 g of the title compound.

Step 3E

3-Carboxymethyl-azetidine-1-carboxylic Acid Benzyl Ester

A solution of 3-ethoxycarbonylmethylazetidine-1-carboxylic acid benzylester (0.30 g, 1.1 mmol), and lithium hydroxide monohydrate (0.091 g,2.2 mmol) in TBF (3 mL) and water (1 mL) was stirred under argon at roomtemperature for 1 hr. After acidification with 2 M aqueous hydrochloricacid (10 mL), the reaction mixture was extracted with ethyl acetate(3×20 ml). The combined ethyl acetate layers were washed with water (10mL), brine (10 mL), dried over anhydrous sodium sulfate, filtered, andconcentrated to give 0.27 g of the title compound.

EXAMPLE 4

(S)-1-Phenyl-N-{4-[2-aminomethyl-3-methyl-5-(3-trifluoromethylphenyl)-3H-imidazol-4-yl]-pyridin-2-yl}-ethylamine

The compound was prepared as described in example 1, replacing3-(benzyloxycarbonylamino)propanoic acid with N-Cbz-glycine in step 1E.

¹H NMR (300 MHz, CD₃OD) d 8.01 (d, J=5.4 Hz, 1H, Pyr), 7.72 (s, 1H, Ar),7.52 (d, J=7.6Hz, 1H,Ar),7.47 (d, J=7.8 Hz, 1H, Ar),7.37 (t, J=7.6 Hz,1H, Ar), 7.32-7.22 (m, 4H, Ar), 7.16 (t, J=7.1 Hz, 1H, Ar), 6.46 (dxd,J=5.4 & 1.5 Hz, 1H, Pyr), 6.35 (s, 1H, Pyr), 4.82 (q, J=6.8 Hz, 1H,PhCHCH₃), 3.96 (s, 2H, CCH₂NH₂), 3.40 (s, 3H, CH₃N), 1.48 (d, J=6.8 Hz,3H, PhCHCH₃); MS (FAB) Calcd for C₂₅H₂₄F₃N₅ (M+H⁺) 451, found 452

EXAMPLE 5

(S)-1-Phenyl-N-{4-[2-(dimethylamino)methyl-3-methyl-5-(3-trifluoromethyl-phenyl)-3H-imidazol-4-yl]-pyridin-2-yl}-ethylamine

The compound was prepared as described in example 1, replacing3-(benzyloxycarbonylamino)propanoic acid with3-carboxymethyl-azetidine-1-carboxylic acid benzyl ester in step 1E.

¹H NMR (300 MHz, CD₃OD) d 8.00 (d, J=5.4 Hz, 1H, Pyr), 7.71 (s, 1H, Ar),7.51 (d, J=7.8 Hz, 1H, Ar), 7.47 (d, J=7.8 Hz, 1H, Ar), 7.37 (t, J=7.8Hz, 1H, Ar), 7.32-7.22 (m, 4H, Ar), 7.16 (t, J=7.1 Hz, 1H, Ar), 6.46(dxd, J=5.4 & 1.5 Hz, 1H, Pyr), 6.37 (s, 1H, Pyr), 4.83 (q, J=7.1 Hz,1H, PhCHCH₃), 3.61 (s, 2H, CCH₂N), 3.45 (s, 3H, CH₃N), 2.31 (s, 6H,N(CH₃)₂), 1.48 (d, J=6.8 Hz, 3H, PhCHCH₃); MS (FAB) Calcd for C₂₇H₂₈F₃N₅(M+H⁺) 479.2, found 480.3

EXAMPLE 6

(S)-1-Phenyl-N-{4-[2-(3-dimethylamino-propyl)-3-methyl-5-(3-trifluoromethylphenyl)-3H-imidazol-4-yl]-pyridin-2-yl}-ethylamine

The compound was prepared as described in example 1, replacing3-(benzyloxycarbonylamino)propanoic acid with N-Cbz-GABA in step 1E.Then a mixture of(S)-1-phenyl-N-{4-[2-(3-amino-propyl)-3-methyl-5-(3-trifluoromethylphenyl)-3H-imidazol-4-yl]-pyridin-2-yl}-ethylamine(0.10 g, 0.21 mmol), formaldehyde (0.10 g, 1.25 mmol), and palladium onactivated carbon (10%) (0.03 g) in methanol (5 mL) was vigorouslystirred under 1 atm hydrogen for 6 hrs. The reaction mixture was thenfiltered, and concentrated. The residue was chromatographed on silicagel, eluting with 90:10:1 CH₂Cl₂:MeOH:NH₄OH to give 0.087 g of the titlecompound.

¹H NMR (300 MHz, CD₃OD) d 8.00 (d, J=5.4 Hz, 1H, Pyr), 7.68 (s, 1H, Ar),7.50 (d, J=7.5 Hz, 1H, Ar), 7.47 (d, J=8.6 Hz, 1H, Ar), 7.38 (t, J=7.7Hz, 1H, Ar), 7.32-7.23 (m, 4H, Ar), 7.16 (t, J=6.9 Hz, 1H, Ar), 6.46(dxd, J=5.4 & 1.5 Hz, 1H, Pyr), 6.34 (s, 1H, Pyr), 4.81 (q, J=6.8 Hz,1H, PhCHCH₃), 3.34 (s, 3H, CH₃N), 2.83 (t, J=7.7 Hz, 2H, CH₂CH₂N), 2.55(t, J=7.7 Hz, 2H, CCH₂CH₂), 2.35 (s, 6H N(CH₃)2), 1.97 (quin, J=7.5 Hz,2H, CH₂CH₂CH₂), 1.48 (d, J=6.8 Hz, 3H, PhCHCH₃); MS (FAB) Calcd forC₂₉H₃₂F₃N₅ (M+H⁺) 507.3, found 508.4

EXAMPLE 7

(S)-1-Phenyl-N-{4-[2-(3-amino-propyl)-3-methyl-5-(3-trifluoromethyl-phenyl)-3H-imidazol-4-yl]-pyridin-2-yl}-ethylamine

The compound was prepared as described in example 1, replacing3-(benzyloxycarbonylamino)propanoic acid with N-Cbz-GABA in step 1E.

1H NMR (300 MHz, CD₃OD) d 8.00 (d, J=5.1 Hz, 1H, Pyr), 7.69 (s, 1H, Ar),7.49 (d, J=7.6 Hz, 1H, Ar), 7.47 (d, J=5.9 Hz, 1H, Ar), 7.37 (t, J=7.8Hz, 1H, Ar), 7.32-7.22(m,4H, Ar),7.16 (t,J=6.9 Hz, 1H, Ar), 6.46 (dxd,J=5.4 & 1.5 Hz, 1H, Pyr), 6.34 (s, 1H, Pyr), 4.81 (q, J=6.8 Hz, 1H,PhCHCH₃), 3.35 (s, 3H, CH₃N), 2.94 (t, J=7.2 Hz, 2H, CH₂CH₂NH₂), 2.89(t, J=7.3 Hz, 2H, CCH₂CH₂), 2.04 (quin, J=7.3 Hz, 2H, CH₂CH₂CH₂), 1.49(d, J=6.8 Hz, 3H, PhCHCH₃); MS (FAB) Calcd for C₂₇H₂₈F₃N₅ (M+H⁺) 479.2,found 480.3

EXAMPLE 8

{4-[3-methyl-2-(morpholin-4-ylmethyl)-5-(3-trifluoromethylphenyl)-3H-imidazol-4-yl]-pyridin-2-yl}-(1-phenylethyl)-amine

The title compound was prepared as described in example 1, replacing3-(benzyloxycarbonylamino)propanoic acid with morpholin-4-yl-acetic acid(from comercially available methyl morpholinoacetate by aqueous acidhydrolysis (6N HCl reflux 2 h)) in step 1E and omission of deprotectionstep 1H.

¹H NMR (300 MHz, CD₃OD) d 8.12 (d, J=5.4 Hz, 1H, Pyr), 7.79 (s, 1H, Ar),7.47 (d, J=8.0 Hz, 1H, Ar), 7.39 (d, J=8.0 Hz, 1H, Ar), 7.25 (m, 6H,Ar), 6.47 (d, J=5.4 Hz, 1H, Pyr), 6.07 (s, 1H, Pyr), 5.44 (d, J=5.6, 1H,NH), 4.62 (m, 1H, PhCHCH₃), 3.70 (m, 4H, CH₂), 3.64 (s, 2H, CH₂), 3.26(s, 3H, CH₃), 2.51 (m, 4H, CH₂), 1.54 (d, J=6.0 Hz, 3H, PhCHCH₃). MS(FAB) Calcd for C₂₉H₃₀F₃N₅O (M+H⁺) 522.2, found 522.2

EXAMPLE 9

(S)-1-Phenyl-N-{4-[2-(3-methylaminopropyl)-3-methyl-5-(3-trifluoromethylphenyl)-3H-midazol-4-yl]-pyridin-2-yl}-ethylamine

The compound was prepared as described in example 7 and then methylatedas shown in step 1D.

¹H NMR (300 MHz, CD₃OD) d 8.00 (d, J=5.4 Hz, 1H, Pyr), 7.69 (s, 1H, Ar),7.49 (d, J=7.3 Hz, 1H, Ar), 7.47 (d,J=6.4 Hz, 1H, Ar), 7.37 (t, J=7.7Hz, 1H, Ar), 7.32-7.23 (m, 4H, Ar), 7.16 (t, J=6.9 Hz, 1H, Ar), 6.46(dxd, J=5.4 & 1.5 Hz, 1H, Pyr), 6.34 (s, 1H, Pyr), 4.81 (q, J=6.4 Hz,1H, PhCHCH₃), 3.36 (s, 3H, CH₃N), 2.87 (t, J=7.5 Hz, 2H, CH2CH₂NH), 2.81(t, J=7.3 Hz, 2H, CCH₂CH₂), 2.49 (s, 3H, NHCH₃), 2.00 (quin, J=7.3 Hz,2H, CH₂CH₂CH₂), 1.48 (d, J=6.8 Hz, 3H, PhCHCH₃); MS (FAB) Calcd for C₂₈H₃₀F₃N₅ (M+H⁺) 493.2, found 494.3

EXAMPLE 10

(S)-1-Phenyl-N-{4-[2-(2-dimethylaminoethyl)-3-methyl-5-(3-trifluoromethylphenyl)-3H-imidazol-4yl]-pyridin-2-yl}-ethylamine

The compound was prepared as described in example 1 then methylated asdescribed in example 6.

¹H NMR (300 MHz, CD₃OD) d 8.00 (d, J=5.1 Hz, 1H, Pyr), 7.69 (s, 1H, Ar),7.50 (d, J=7.6 Hz, 1H, Ar), 7.46 (d, J=7.8 Hz, 1H, Ar), 7.36 (t, J=7.7Hz, 1H, Ar), 7.31-7.22 (m, 4H, Ar), 7.16 (t, J=6.9 Hz, 1H, Ar), 6.46(dxd, J=5.1 & 1.5 Hz, 1H, Pyr), 6.34 (s, 1H, Pyr), 4.81 (q, J=6.8 Hz,1H, PhCHCH₃), 3.36 (s, 3H, CH₃N), 2.99 (t, J=7.6 Hz, 2H, CH₂CH₂N), 2.78(t, J=7.6 Hz, 2H, CCH₂CH₂), 2.37 (s, 6H, N(CH₃)₂), 1.48 (d, J=6.8 Hz,3H, PhCHCH₃); MS (FAB) Calcd for C₂₈H₃₀F₃N₅ (M+H⁺) 493.2, found 494.3

The ability of compounds of the present invention to inhibit thesynthesis or the activity of cytokines can be demonstrated using thefollowing in vitro assays.

BIOLOGICAL ASSAYS

Lipopolysaccharide Mediated Production of Cytokines

Human peripheral blood mononuclear cells (PBMC) are isolated from freshhuman blood according to the procedure of Chin and Kostura, J. Immunol.151, 5574-5585 (1993). Whole blood is collected by sterile venipunctureinto 60 mL syringes coated with 1.0 mL of sodium-heparin (Upjohn, 1000U/mL) and diluted 1:1 in Hanks Balanced Salt Solution (Gibco). Theerythrocytes are separated from the PBMC's by centrifugation on aFicoll-Hypaque lymphocyte separation media. The PBMC's are washed threetimes in Hanks Balanced Salt Solution and then resuspended to a finalconcentration of 2×10⁶ cell/mL in RPMI containing 10% fresh autologoushuman serum, penicillin streptomycin (10 U/mL) and 0.05% DMSO.Lipopolysaccharide (Salmonella type Re545; Sigma Chemicals) is added tothe cells to a final concentration of 100 ng/mL. An aliquot (0.1 mL) ofthe cells is quickly dispensed into each well of a 96 well platecontaining 0.1 mL of the test compound, at the appropriate dilution, andare incubated for 24 hours. at 37° C. in 5% CO2. At the end of theculture period, cell culture supernatants are assayed for IL-1β, TNF-α,IL-6 and PGE2 production using specific ELISA.

IL-1 Mediated Cytokine Production

Human peripheral blood mononuclear cells are isolated from fresh humanblood according to the procedure of Chin and Kostura, J. Immunol. 151,5574-5585 (1993). Whole blood is collected by sterile venipuncture into60 mL syringes coated with 1.0 mL of sodium-heparin (Upjohn, 1000 U/mL)and diluted 1:1 in Hanks Balanced Salt Solution (Gibco). Theerythrocytes are separated from the PBMC's by centrifugation on aFicoll-Hypaque lymphocyte separation media. The PBMC's are washed threetimes in Hanks Balanced Salt Solution and then resuspended to a finalconcentration of 2×10⁶ cell/mL in RPMI containing 10% fresh autologoushuman serum, penicillin strepto-mycin (10 U/ml) and 0.05% DMSO.Endotoxin free recombinant human IL-1b is then added to a finalconcentration of 50 pMolar. An aliquot (0.1 mL) of the cells is quicklydispensed into each well of a 96 well plate containing 0.1 mL of thecompound at the appropriate dilution. and are incubated for 24 hours. at37° C. in 5% CO2. At the end of the culture period, cell culturesupernatants are assayed for TNF-a, IL-6 and PGE2 synthesis usingspecific ELISA.

Determination of IL-1β, TNF-α, IL-6 and Prostanoid Production From LPSor IL-1 Stimulated PBMC's

IL-1β ELISA

Human IL-1β can be detected in cell-culture supernatants or whole bloodwith the following specific trapping ELISA. Ninety-six well plasticplates (Immulon 4; Dynatech) are coated for 12 hours at 4° C. with 1mg/mL protein-A affinity chromatography purified mouse anti-human IL-1βmonoclonal antibody (purchased as an ascites preparation from LAOEnterprise, Gaithersburg Md.) diluted in Dulbecco's phosphate-bufferedsaline (—MgCl₂, —CaCl₂). The plates are washed with PBS-Tween(Kirkegaard and Perry) then blocked with 1% BSA diluent and blockingsolution (Kirkegaard and Perry) for 60 minutes at room temperaturefollowed by washing with PBS Tween. IL-1β standards are prepared frompurified recombinant IL-1β produced from E. coli. The highestconcentration begins at 10 ng/mL followed by 11 two-fold serialdilutions. For detection of IL-1β from cell culture supernatants orblood plasma, 10-25 mL of supernatant is added to each test well with75-90 mL of PBS Tween. Samples are incubated at room temperature for 2hours then washed 6 times with PBS Tween on an automated plate washer(Dennly). Rabbit anti-human IL-1β polyclonal antisera diluted 1:500 inPBS-Tween is added to the plate and incubated for 1 hour at roomtemperature followed by six washes with PBS-Tween. Detection of boundrabbit anti-IL-1β IgG is accomplished with Fab′ fragments of Goatanti-rabbit IgG-horseradish peroxidase conjugate (Accurate Scientific)diluted 1:10,000 in PBS-Tween. Peroxidase activity was determined usingTMB peroxidase substrate kit (Kirkegaard and Perry) with quantitation ofcolor intensity on a 96-well plate Molecular Devices spectrophotometerset to determine absorbance at 450 nM. Samples are evaluated using astandard curve of absorbance versus concentration. Four-parameterlogistics analysis generally is used to fit data and obtainconcentrations of unknown compounds.

TNF-α ELISA

Immulon 4 (Dynatech) 96-well plastic plates are coated with a 0.5 mg/mLsolution of mouse anti-human TNF-a monoclonal antibody. The secondaryantibody is a 1:2500 dilution of a rabbit anti-human TNF-α polyclonalserum purchased from Genzyme. All other operations are identical tothose described above for IL-1β. The standards are prepared inPBS-Tween+10% FBS or HS. Eleven 2 fold dilutions are made beginning at20 ng/mL TNF-α.

IL-6 ELISA

Levels of secreted human IL-6 are also determined by specific trappingELISA as described previously in Chin and Kostura, J. Immunol. 151,5574-5585 (1993). (Dynatech) ELISA plates are coated with mouseanti-human IL-6 monoclonal antibody diluted to 0.5 mg/mL in PBS. Thesecondary antibody, a rabbit anti-human IL-6 polyclonal antiserum, isdiluted 1:5000 with PBS-Tween. All other operations are identical tothose described above for IL-1b. The standards are prepared inPBS-Tween+10% FBS or HS. Eleven 2 fold dilutions are made beginning at50 ng/mL IL-6.

PGE₂ Production

Prostaglandin E2 is detected in cell culture supernatants from LPS orIL-1 stimulated PBMC's using a commercially available enzymeimmunoassay. The assay purchased from the Cayman Chemical (Cataloguenumber 514010) and is run exactly according to the manufacturersinstructions.

Interleukin8 (IL-8)

The present compounds can also be assayed for IL-8 inhibitory activityas discussed below. Primary human umbilical cord endothelial cells(HUVEC) (Cell Systems, Kirland, Wash.) are maintained in culture mediumsupplemented with 15% fetal bovine serum and 1% CS-HBGF consisting ofaFGF and heparin. The cells are then diluted 20-fold before being plated(250 μl) into gelatin coated 96-well plates. Prior to use, culturemedium is replaced with fresh medium (200 μl). Buffer or test compound(25 μl, at appropriate concentrations) is then added to each well inquadruplicate wells and the plates incubated for 6 h in a humidifiedincubator at 37° C. in an atmosphere of 5% CO₂. At the end of theincubation period, supernatant is removed and assayed for IL-8concentration using an IL-8 ELISA kit obtained from R&D Systems(Minneapolis, Minn.). All data is presented as mean value (ng/mL) ofmultiple samples based on the standard curve. IC50 values whereappropriate are generated by non-linear regression analysis.

What is claimed is:
 1. A compound of the formula

wherein X is C₁-C₆ alkyl, said alkyl group optionally substituted by 1-3groups selected from halogen, hydroxy, CF₃, NH₂ and NO₂; or aheterocyclic group connected to the imidazole ring by a direct bond orby C₁-C₆ alkyl; R₁ and R₂ are independently hydrogen or C₁-C₆ alkyl,said alkyl group optionally substituted by 1-3 groups selected fromhalogen, hydroxy, CF₃, NH₂ and NO₂; or R₁ and R₂ taken together with thenitrogen atom represent an optionally substituted 4 to 10 memberednon-aromatic heterocyclic ring containing at least one N atom, andoptionally containing 1-2 additional N atoms and 0-1 O or S atom; or R₂and X taken together represent an optionally substituted 4 to 10membered non-aromatic heterocyclic ring containing at least one N atom,and optionally containing 1-2 additional N atoms and 0-1 O or S atoms; Qis CH or N; R₃ is hydrogen or NH(C₁-C₆ alkyl)aryl; R₄, R₅ and R₆independently represent a member selected from the group consisting ofhydrogen, halo, hydroxy, CF₃, NH₂, NO₂, C₁-C₆ alkyl, said alkyl groupoptionally substituted by 1-3 groups selected from halogen, hydroxy,CF₃, NH₂ and NO₂, C₁-C₆ alkoxy, said alkoxy group optionally substitutedby 1-3 groups selected from halogen, hydroxy, CF₃, NH₂ and NO₂; C₃-C₈cycloalkyl, said cycloalkyl group optionally substituted by 1-3 groupsselected from halogen, hydroxy, CF₃, NH₂ and NO₂ or aryl, said arylgroup optionally substituted by 1-3 groups selected from halogen,hydroxy, CF₃, NH₂ and NO₂; R₇ is hydrogen or C₁-C₆ alkyl, said alkylgroup optionally substituted by 1-3 groups selected from halogen,hydroxy, CF₃, NH₂ and NO₂; or a pharmaceutically acceptable additionsalt and/or hydrate thereof, or where applicable, a geometric or opticalisomer or racemic mixture thereof.
 2. The compound in accordance withclaim 1 of the formula

or a pharmaceutically acceptable salt thereof, wherein: Q is CH; X isC₁-C₆ alkyl; or R₂ and X taken together represent an azetidine group;R₄, R₅ and R₆ are independently hydrogen or CF₃; R₇ is CH₃; R₁ and R₂are independently hydrogen or C₁-C₆ alkyl; or R₁ and R₂ taken togetherwith the nitrogen atom form a morpholine group.
 3. A compound of theformula:

or a pharmaceutically acceptable addition salt and/or hydrate thereof,or where applicable, a geometric or optical isomer or racemic mixturethereof.
 4. A pharmaceutical composition which is comprised of acompound in accordance with claim 1 in combination with apharmaceutically acceptable carrier.
 5. A pharmaceutical compositionwhich is produced by combining a compound in accordance with claim 1 anda pharmaceutically acceptable carrier.
 6. A method of treating acytokine mediated disease in a mammal, comprising administering to amammalian patient in need of such treatment an amount of a compound asdescribed in claim 1 in an amount which is effective to treat saidcytokine mediated disease.
 7. A method of treating inflammation in amammalian patient in need of such treatment, which is comprised ofadministering to said patient an anti-inflammatory effective amount of acompound as described in claim
 1. 8. A method in accordance with claim 6wherein the cytokine mediated disease is rheumatoid arthritis,osteoarthritis, endotoxemia, toxic shock syndrome, inflammatory boweldisease, tuberculosis, atherosclerosis, muscle degeneration, cachexia,psoriatic arthritis, Reiter's syndrome, rheumatoid arthritis, gout,traumatic arthritis, rubella arthritis or acute synovitis.
 9. A methodin accordance with claim 6 wherein the cytokine mediated disease isrheumatoid arthritis, rheumatoid spondylitis, osteoarthritis, goutyarthritis, sepsis, septic shock, endotoxic shock, gram negative sepsis,toxic shock syndrome, adult respiratory distress syndrome, cerebralmalaria, chronic pulmonary inflammatory disease, silicosis, pulmonarysarcosis, bone resorption diseases, reperfusion injury, graft v. hostrejection, allograft rejection, fever, myalgia due to infection,cachexia secondary to infection or malignancy, cachexia secondary toacquired immune deficiency syndrome (AIDS), AIDS related complex (ARC),keloid formation, scar tissue formation, Crohn's disease, ulcerativecolitis or pyresis.
 10. A method of treating osteoporosis in a mammalianpatient in need of such treatment, which is comprised of administeringto said patient an amount of a compound as described in claim 1 which iseffective to treat osteoporosis.
 11. A method of treating boneresorption in a mammalian patient in need of such treatment, which iscomprised of administering to said patient an amount of a compound asdescribed in claim 1 which is effective to treat bone resorption.
 12. Amethod of treating Crohn's disease in a mammalian patient in need ofsuch treatment which is comprised of administering to said patient anamount of a compound as described in claim 1 which is effective to treatCrohn's disease.
 13. A process for making a pharmaceutical compositioncomprising combining a compound of claim 1 and a pharmaceuticallyacceptable carrier.